The phenomena of dielectrophoresis, alternating current (AC) electrothermal effect and AC electroosmosis, collectively referred to as alternating current (AC) electrokinetics (ACEK), are now being used to manipulate and separate particles on a cellular scale. Dielectrophoresis (DEP) involves the suspension of a dielectric particle in a non-uniform electric field. As will be discussed further herein, capacitive and impedance changes may be recognized from, for example, a two (or more) electrode array coated with a molecular probe (such as bacterial antigen) for substances under examination. If a polarized particle is suspended in such a field, an induced dipole will form across the particle and rotate or move in synchrony with the field. Furthermore, as will be depicted and discussed herein, the AC electrothermal and AC electroosmosis phenomena or effects will induce microscale flows around the electrodes, convecting particles/colloids/macromolecules to the electrodes for detection.
Interdigitated micro-electrodes or two closely spaced parallel plates are known and described, for example, in Capacitive Microsystems for Biological Sensing, V. Tsouti et al., Biosensors and Biolelectronics, 27, (2011), pp. 1-11. In simplified form, electrodes of a capacitance-type sensor may comprise two closely spaced parallel plates having particular spacing and thickness. A parallel connection of capacitors having two electrodes may be formed. It is well known that the sum of the individual capacitors in parallel comprises the capacitance of the parallel capacitors. While described as capacitors, no capacitor exhibits perfect capacitance without resistive and inductive components to create an impedance. Yet, the resistive and inductive components of such capacitive microsystems are less indicative of surface binding compared to the capacitive component. Such biosensors have been particularly developed and utilized, for example, in the detection of Escherichia coli and salmonella. Another electrode array is known for prostate specific antigen (PSA) testing for prostate cancer. Yet another prototype test integrated circuit has been developed for certain protein detection.
Johne's disease is caused by bacteria known as Mycobacterium avium subspecies paratuberculosis. Johne's disease affects wildlife and livestock. In livestock such as cattle or dairy cows, the disease causes reduction of milk production (dairy cows), weight loss and premature culling of clinically affected animals. In the United States alone, Johne's disease has been found in 68% of dairy herds and causes an estimated annual loss of $220 million to the US dairy industry alone. Johne's disease is currently diagnosed in diagnostic laboratories using immunoassay or enzyme-linked imnmunosorbent assay (ELISA) or pathogen detection methods (bacterial culture or PCR indicative of infection or contamination).
Mycobacterium bovis causes bovine tuberculosis both in animals and humans. Despite progress towards eradication of bovine tuberculosis from US livestock, states like Michigan and Minnesota continue to struggle with bovine tuberculosis in their wildlife and cattle operation. Mandatory testing of cattle costs $3.25 million per year in Minnesota alone. In the U.S., incidences of bovine tuberculosis cost more than $40 million in 2008-2009 for testing and treatment. Bovine tuberculosis in wild animals is currently tested by postmortem examination of gross lesion, bacterial culture, and skin test.
Human tuberculosis, caused by Mycobacterium tuberculosis, occurs in more than ten million people and, worldwide, is estimated to be responsible for the death of two million people annually. It is estimated that over one billion dollars is spent on diagnosis and evaluation of human tuberculosis worldwide each year. Human tuberculosis is currently diagnosed by radiographic imaging (conventional chest x-ray), smear microscopy, bacterial culture, or a tuberculin skin test.
Mastitis is a disease that results in inflammation of the mammary gland that is mostly caused by bacterial infections. The disease is the most common cause of death in adult dairy cattle. Indeed, it is estimated that 38% of all cows are affected with mastitis. Mastitis causes an estimated 1.7-2.0 billion USD annual economic loss to the US dairy industry. Worldwide, it is the most costly disease affecting the dairy industry, incurring economic losses estimated at $50 billion/year (˜£31 billion/year). Escherichia coli and Streptococcus uberis are common causative agents of bovine mastitis and are responsible for about 18% and 5% of the disease, respectively. Bacterial counts in milk of mastitis cow can reach 107 bacteria/ml. A further indication of mastitis in lactating animals is somatic white blood cell count which can be determined by mixing infected cow milk with a reagent and the amount of gel formed indicates a count of somatic cells and so an indication of mastitis. Detection and identification of the bacteria in fresh milk are critically important for treatment and control of the disease in dairy farms.
From U.S. Pat. Nos. 7,517,955 and 7,812,147 assigned to the University of Tennessee Research Foundation, a polypeptide, designated “Streptococcus uberis Adhesion Molecule” or SUAM, was developed by a team comprising Stephen P. Oliver et al. SUAM may be used diagnostically and therapeutically. The patents further describe an immune-fluorescence milk card-test and an agglutination/precipitation test that may be used “cow-side” for diagnosis as well as known ELISA testing which may require hours in a laboratory for results.
In the home and in the field, it would be beneficial if a laboratory on an integrated circuit (chip), as has been developed for other diseases, and related methodology may be available for rapid testing of wildlife, livestock and humans for diseases and physiological conditions such as bacterial diseases including tuberculosis, Johne's disease, mastitis and instances of heart attack among other diagnosis.
D-Dimer is an indicator of the degradation of a clot and, hence, is a predictor or indicator of a pulmonary embolism, deep venous thrombosis and the like. Clots are often fatal for example a clot that may form in a vein and return to the heart. It is desirable to have a lab-on-a-chip test for the detection of D-dimer.
High/low sugar content, for example, glucose of the blood and other bodily fluids is an indicator of hyper or hypo glycemia among other predictors of sugar related disease. A lab-on-a chip test for sugar content may help patients and doctors determine such sugar related ease immediately and compete with existing methodology. Moreover, a possible industrial or commercial application is, for example, to test sugar content in beer.
Small molecule detection generally relates to any small molecule that may be a predictor of a disease of a condition. Specifically, it may, for example, be desirable to test for progesterone as an example of a condition such as pregnancy.
Enzymes are complexes produced, for example, in living cells of human organs or skeletal structures. Consequently, while ELISA testing is available, there is a need for a simple lab-on-a-chip test for enzyme level that can be an organ disease marker and accomplish in minutes what ELISA may require a formal laboratory and days to obtain results.
Another potential lab-on-a-chip application is in the testing of well water for coliform or E. coli bacteria in water rather than wait for a culture of other slow laboratory means for testing known in the art. Another bacteria requiring swabbing and testing is Streptococcus which is an indicator, for example, of strep throat.
Given the foregoing, what are needed are methods and related laboratory-on-a-chip apparatus that may provide for detection of bacterial and other infectious diseases, conditions via biomarkers or even for use in commercial applications, for example, using AC electrokenetics phenomena.